This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The functional properties of rhesus cytomegalovirus (rhCMV)-specific CD8-positive T lymphocytes with regards to cytotoxicity and interferon (IFN)-gamma secretion were investigated in six asymptomatic CMV-seropositive rhesus macaques. CD8-positive T lymphocytes recognizing ten rhCMV epitopes, one of which was identical in three animals, were analyzed. Optimal flow-cytometric conditions to detect lysosomal-associated membrane proteins (LAMPs) on the surface of recently degranulated cells in rhesus macaques were established and revealed that unlike humans, the rhesus macaque LAMP-1 protein CD107a underwent little or no endocytosis. Following in vitro stimulation, rhCMV-specific CD8-positive T lymphocytes were heterogeneous with regards to time to reach peak degranulation, kinetics of IFN-gamma secretion relative to degranulation, and composition of cells positive for CD107a and/or IFN-gamma. Responder CD8-positive T lymphocytes that underwent degranulation without IFN-gamma production were enriched for effectors (CD28-negativeCD45RA-positive), and had significantly lower frequencies of effector memory (CD28-negativeCD45RA-negative) cells compared to the IFN-gamma-secreting cells that did or did not undergo degranulation. The presence of phenotypically distinct populations of rhCMV-specific CD8-positive T lymphocytes with variable cytolytic and IFN-gamma-secreting ability may determine their relative ability to control CMV replication. These data underscore the necessity of using more than one effector function for evaluating virus-specific CD8-positive T lymphocytes. AIDS related.